Accumulating evidence suggests that dysregulation of marginal zone macrophages (MZMs) of the spleen plays a role in the development of systemic autoimmune disease include systemic lupus erythematosus (SLE). The MZMs play an important role in trapping, processing, and inducing tolerance to apoptotic cell (AC) antigens. The molecular mechanisms underlying dysregulation of tolerogenic MZM responses in lupus and the signals that trigger this dysregulation have not been elucidated. The proposed studies test the hypotheses that: (1) A common mechanism underlying dysregulation of tolerogenic MZM responses in lupus is an alteration in the expression of a mechanosensing protein megakaryocytic leukemia-1 (MKL1) and associated filamentous actin assembly in MZMs that affects the ability of the MZMs to take up and maintain tolerance to apoptotic cell antigens (AC-Ags); (2) Alterations in expression of MKL1 and disruption of F-actin assembly can be induced by events that are characteristic of SLE. They can be induced by loss of signaling through the lymphotoxin- receptor (LTR) on the MZMs resulting from type I interferon (IFN) induced mislocalization of MZ B cells, which express membrane lymphotoxin (mLT), from the MZ to the follicle. Notably, the interferon signature is a hallmark of SLE in humans and deficiency of Type 1 IFN signals in BXD2- Ifnr-/- mice abrogates many of the abnormal immunologic MZM phenotypes as well as the production of autoantibodies and symptomatic lupus in these mice. The alterations in MKL1 and actin assembly also can be induced by chronic exposure to AC debris and immune complexes (ICs) through mechanosensing. These hypotheses will be tested using multiple mouse models with targeted disruption of pertinent pathways made available through extensive collaborations, pharmacological disruption, and state-of-the-art confocal microscopic analysis. Aim 1 will determine if IFN- induced follicular translocation of mLT+ MZ B cells leads to loss of MZM tolerance of ACs and ICs. In addition, BXD2 and B6.TC autoimmune mice will be used to determine if an MZM defect is a primary, early defect in lupus. Aim 2 will focus on the LT R downstream signaling of MKL1 pathway and test in vivo and in vitro if the LTR/MLK1/actin polymerization pathways play a role in maintenance of the survival of tolerogenic MZMs. Frozen sections from SLE and normal human spleen will be used to determine if loss of MARCO+ MZMs in the SLE spleen is associated with high numbers of type 1-IFN producing pDCs, follicular translocation of LT+ B cells, and dysregulation of MKL1/actin polymerization. Clinical Relevance. The studies will provide a unified model of lupus, indicate a critical novel pathogenic mechanism of type I IFNs, and identify a new molecular pathway underlying regulation of tolerogenic macrophages in SLE thereby suggesting novel candidate therapeutic targets.